2.1 Amyotrophic Lateral Sclerosis
Amyotrophic lateral sclerosis (ALS) is a relentless fatal paralytic disorder confined to the voluntary motor system [37]. The onset of disease is usually in the fourth or fifth decade of life. Common clinical features of ALS include muscle weakness, fasciculations, brisk (or depressed) reflexes, and extensor plantar responses. Although motor deficit usually predominates in the limbs, bulbar enervation can be severely involved, sometimes early in the course of the disease, leading to atrophy of the tongue, dysphagia, and dysarthria. Other cranial nerves (e.g., occulomotor nerves) are usually spared, unless the patient survives beyond respiratory failure [38]; this latter is due to respiratory muscle paralysis that occurs frequently in advanced cases. The disease progresses rapidly, with a mean survival of 3 years. Pathologically, ALS is characterized mainly by a loss of upper motor neurons, lower motor neurons, or both [39]. To date, only a few approved treatments, such as mechanical ventilation and riluzole, do prolong survival in ALS patients to some extent. However, the development of more effective neuroprotective therapies is impeded by our limited knowledge of the actual mechanisms by which motor neurons die in ALS, and of how the disease propagates and progresses.
ALS usually arises as a sporadic condition in the absence of any apparent genetic linkage, but occasionally (in about 10 percent of cases) the disease is inherited [40]. The majority of familial forms of ALS are transmitted as autosomal dominant traits and are clinically and pathologically almost indistinguishable from sporadic ALS; however, familial ALS tends to have an earlier age of onset, a more rapid course, and a survival after diagnosis of only 1.5 years. Approximately 20 percent of familial ALS cases are linked to mutations in the gene encoding for the cytosolic enzyme SOD1 [40]. To date, more than 120 mutations in SOD1 have been identified in familial ALS families. Many of these mutations lead to the substitution of an amino acid, several of which, such as the glycine-to-alanine substitution at position 93 (G93A) and the glycine-to-arginine substitution at position 37 (G37R), are similar to wild-type SOD1 with respect to their stability, metal coordination properties and homospecific catalytic activity, whereas several others, such as the glycine-to-arginine substitution at position 85 (G85R), exhibit poor stability and low catalytic activity [41].
For more than a decade, the lion's share of attention in ALS research has been paid to mutant SOD1. The rationale for studying so avidly this rare form of ALS rests on the expectation that the phenotypic similarity between the genetic and sporadic forms of the disease indicates that they share important pathogenic mechanisms and, consequently, that information generated by studying mutant SOD1 will help focus research on key cellular and molecular mechanisms.
SOD1 is an abundant, ubiquitously expressed cytosolic enzyme whose known activity is to dismutate superoxide to hydrogen peroxide. Although SOD is thought to be essential for living organisms [42], mutant mice deficient in this enzyme thrive normally and do not develop a ALS phenotype [43]. Conversely, Tg (Tg) rodents expressing either catalytically active SOD1 mutants [3; 4] or catalytically inactive SOD1 mutants [5; 44] recapitulate the clinical and the neuropathological hallmarks of ALS. Transgenic (Tg) mice expressing high levels of wild-type human SOD1 are healthy [3]. Taken together, these results argue for mutant SOD1 causing motor neuron degeneration, not via a loss-of-, but rather via a gain-of-function effect. However, despite intense research efforts, the nature of the adverse property manifested by mutant SOD1 remains elusive. To date, it has been proposed that mutant SOD1 cytotoxicity involves different mechanisms including oxidative stress [45, 46], protein aggregation [14], aberrant protein-protein interactions [48], decreased binding affinity for zinc [49], mitochondrial dysfunction [50], and apoptosis [9, 51], none of which are mutually exclusive,
Aside from the mechanism of toxicity, the mutant SOD1 cellular site of action is also a source of discussion. For instance, several recent studies support the notion that mutant SOD1 in both motor neurons and non-neuronal cells contribute to the disease process in vivo [52, 53, 8]. Indeed, it has been shown that both the selective lowering of mutant SOD1 in either motor neurons or in glial cells such as microglia by a Cre-Lox system prolongs survival in Tg SOD1G37R mice compared to their germline littermates [8]. Similarly, PU.1-deficient mice carrying the SOD1G93A mutation, and in which the lack of microglia is corrected by transplantation of wild-type bone marrow cells, have a longer lifespan than those transplanted with SOD1G93A bone marrow cells [53]. Furthermore, studies in chimeric mice [20] composed of cells expressing either wild-type or mutant SOD1 observed that: (i) a greater fraction of mutant motor neurons survived when surrounded by wild-type cells; and, (ii) wild-type motor neurons surrounded by non-neuronal cells expressing mutant SOD1 did acquire ubiquitin-positive protein aggregates, a sign of neuronal damage in this ALS model [9]. In a model system of a different form of neurodegeneration, an effect of supporting cells on nearby neurons has also been observed: it has been shown that expression of mutant ataxin-7 in astrocytes causes degeneration of wild-type Purkinje cells in a mouse model of spinocerebellar atrophy [10].
As indicated above, deletion of mutant SOD1 in microglia [22] or absence of microglia expressing mutant SOD1 [53] prolonged survival in Tg mutant SOD1 mice. However, this did not delay the age at onset of symptoms. Furthermore, mutant SOD1 microglia did not induce the death of wild-type motor neurons in vivo [53].
2.2 Spinal Muscular Atrophy
Aside from ALS, there are several other types of prominent motor neuron disease [37]. Among these, spinal muscular atrophy (“SMA”) is the most common fatal neurodegenerative disease of infancy, with an incidence of 1 in 6,000 [54]. This autosomal recessive disease maps to the proximal region of the long arm of chromosome 5, which contains two, almost identical, genes termed “survival of motor neuron” (SMN) genes [55]. In 95 percent of the SMA patients, there is a large-scale deletion of the telomeric copy of SMN, designated SMN1, whereas, in the 5 percent remaining, there is a point mutation in or a short deletion of SMN1 [26]. These observations led to the conclusion that SMN1 is a determining gene in SMA. Conversely, it was found that SMA patients always carry at least one copy of SMN in the form of a centromeric copy, designated SMN2 [54]. This gene, however, is only partially functional, and thus SMN2 is unable to fully compensate for the SMN1 defect [54]. Notably, due to the unstable nature of the genome that contains the SMN genes, SMA patients can carry more than one copy of SMN2 [54] and the greater the number of SMN2 copies, the milder the disease phenotype [56]. For instance, 80 percent of patients with type-I SMA (i.e. severe weakness, profound hypotonia, and a mortality usually due to respiratory failure within the 2 first years of life) carry one or two SMN2 copies; 82 percent of patients with type-II SMA (i.e. disability of later onset, less severe weakness, and survival into adolescence and beyond) carry three SMN2 copies; and, 96 percent of patients with type-III SMA (i.e. onset usually in adolescence or youth adulthood, mild weakness allowing an achievement of ambulation and a normal survival expectancy) carry three or four SMN2 copies [56]. Therefore, SMN2 is not a SMA-causing gene, but it is an important disease modifier, a concept that has been confirmed in engineered mice [57].
While significant strides have been made over the past decade in unraveling the neurobiology of SMA, the function of the SMN protein still remains incompletely elucidated [54]. SMN is ubiquitously expressed and, if knocked out, any cell type would die [58]. This provides compelling evidence that SMN plays a vital role and, by now, it is well established that, through its binding capacity to ribonucleoproteins, SMN is essential to the assembly of the proper Sm core protein to small nuclear RNAs [59]. In addition to its critical role in the biogenesis of the small nuclear ribonucleoproteins, SMN also participates in the maturation of pre-mRNA [60]. However, SMN binds to several other partners [61-64] whose functions are distinct from that of small nuclear ribonucleoprotein biogenesis and pre-mRNA splicing, hence supporting the notion that SMN must be endowed with other important, and for the moment unknown, cellular roles.
Another unsettled question about SMN is: why does a ubiquitously expressed protein cause a motor neuron disease? It is already known that SMN is highly expressed in spinal motor neurons [54] and that lowering SMN selectively in neurons provokes their demise [36]. However, since a low level of SMN is noxious to all cells, one cannot exclude that reduced SMN levels contribute to the disease phenotype by also affecting non-neuronal cells such as glia.